It has been acknowledged that dead bacteria, and bacterial components such as a cell wall skeleton integrant (hereinafter, it is abbreviated as CWS), muramyl dipeptide (MDP), a lipopolysaccaride (LPS), mannan and glucan of bacteria, and a derivative thereof (it may include substances produced by recombinant DNA technique) have an immunopotentiating activity, and exhibits an anti-tumor effect for example in experimental cancer cell lines and in immunotherapy for human cancers. Further, it has been acknowledged that, when formulated into an oil-in-water emulsion by dispersing it in an oil with a dispersing or emulsifying device such as a homogenizer, and besides by emulsifying the dispersion in a surfactant solution, the bacterial components exhibit a better anti-tumor and infection-preventing effect via an immunopotentiating activity (Cancer Res., 33, 2187-2195 (1973), J. Nat. Cancer Inst., 48, 831-835 (1972), J. Bacteriol., 94, 1736-1745 (1967), Gann, 69, 619-626 (1978), J. Bacteriol., 92, 869-879 (1966)).
However, although oil-in-water emulsions as mentioned above have gained a good reputation in utility (Proc. Japan Acad., 70, Ser. B205-209 (1994); Proc. Japan Acad., 74, Ser. B20550-55 (1998)), there have been any problem that the emulsions are prohibited to be constantly, supplied to market since it is difficult, due to the following reasons, that they are commercially produced. The present invention was developed to overcome the various drawbacks that make their commercial production difficult, and resolve the problem as mentioned above. Such drawbacks and means for resolving the same are discussed as shown below.
In general, an oil-in-water emulsion is liable to be largely altered with the time course, resulting in separation between oil and aqueous phases. Thus, to stabilize an emulsion, various measures have been taken such as (1) making particles, (2) reducing the difference in specific gravity between a dispersion medium and a dispersoid, and (3) elevating the viscosity of a dispersion medium with addition of a high-molecular substance, and the like. However, all of them merely extend a period of time required for the alteration, and finally lead to the separation between oil and aqueous phases
Particularly, an oil-in-water emulsion comprising a bacterial component as an active ingredient was shown to be unstable due to the included CWS, thus generating insoluble aggregations in a few days, and therefore, there was no means but the preparation just before use in order to prepare an oil-in-water emulsion. To cope with this point, a sugar alcohol and a saccharide was tried in formulations by lyophilization (Japanese Patent Publication (kokoku) No. 1291/1988). However, the formulations in the state of the art demonstrated large changes in average particle diameter and particle size distribution immediately after lyophilization and as short as one month after the preservation, and therefore, any emulsions have not accomplished significant, improvement in stability that provides practical use.